ENS - Départment de biologie

Marie-Anne Félix, médaille d'argent du CNRS 2012

Marie Embs — 2015-02-25T11:14:44Z

http://www.cnrs.fr/fr/recherche/prix/medaillesargent.htm]

Marie-Anne Félix, médaille d'argent du CNRS 2012

Marie Embs — 2015-02-25T11:14:07Z

http://www.cnrs.fr/fr/recherche/prix/medaillesargent.htm]

Fast multicolor 3D imaging using aberration-corrected multifocus microscopy

Marie Embs — 2015-02-25T10:49:53Z

Conventional acquisition of three-dimensional (3D) microscopy data requires sequential z scanning and is often too slow to capture biological events. We report an aberration-corrected multifocus microscopy method capable of producing an instant focal stack of nine 2D images. Appended to an epifluorescence microscope, the multifocus system enables high-resolution 3D imaging in multiple colors with single-molecule sensitivity, at speeds limited by the camera readout time of a single image.

http://www.nature.com/nmeth/journal/vaop/ncurrent/abs/nmeth.2277.html

Nature Methods (2012) doi:10.1038/nmeth.2277

Sonia Garel, lauréate de l'EMBO Young Investigator 2012

Marie Embs — 2015-02-25T10:49:01Z

http://www.embo.org/news-a-media-centre/press-releases/22-young-group-leaders-recognized-as-embo-young-investigators.html

Antoine Triller, lauréat de l'appel d'offre ERC Advanced Grant 2012

Marie Embs — 2015-02-25T10:48:09Z

Plus d'informations sur le site de l'ERC

CLIP-seq of eIF4AIII reveals transcriptome-wide mapping of the human exon junction complex

Marie Embs — 2015-02-25T10:47:14Z

The exon junction complex (EJC) is a central effector of the fate of mRNAs, linking nuclear processing to mRNA transport, translation and surveillance. However, little is known about its transcriptome-wide targets. We used cross-linking and immunoprecipitation methods coupled to high-throughput sequencing (CLIP-seq) in human cells to identify the binding sites of the DEAD-box helicase eIF4AIII, an EJC core component. CLIP reads form peaks that are located mainly in spliced mRNAs.
Most expressed exons harbor peaks either in the canonical EJC region, located 24 nucleotides upstream of exonic junctions, or in other noncanonical regions. Notably, both of these types of peaks are preferentially associated with unstructured and purine-rich sequences containing the motif GAAGA, which is a potential binding site for EJC-associated factors. Therefore, EJC positions vary spatially and quantitatively between exons. This transcriptome-wide mapping of human eIF4AIII reveals unanticipated aspects of the EJC and broadens its potential impact on post-transcriptional regulation.

Nat Struct Mol Biol. 2012 Oct 21. doi : 10.1038/nsmb.2420. [Epub ahead of print]

Analysis of human samples reveals impaired SHH-dependent cerebellar development in Joubert syndrome/Meckel syndrome

Marie Embs — 2015-02-25T10:46:14Z

Joubert syndrome (JS) and Meckel syndrome (MKS) are pleiotropic ciliopathies characterized by severe defects of the cerebellar vermis, ranging from hypoplasia to aplasia. Interestingly, ciliary conditional mutant mice have a hypoplastic cerebellum in which the proliferation of cerebellar granule cell progenitors (GCPs) in response to Sonic hedgehog (SHH) is severely reduced. This suggests that Shh signaling defects could contribute to the vermis hypoplasia observed in the human syndromes. As existing JS/MKS mutant mouse models suggest apparently contradictory hypotheses on JS/MKS
etiology, we investigated Shh signaling directly on human fetal samples. First, in an examination of human cerebellar development, we linked the rates of GCP proliferation to the different levels and localizations of active Shh signaling and showed that the GCP possessed a primary cilium with CEP290 at its base. Second, we found that the proliferation of GCPs and their response to SHH were severely impaired in the cerebellum of subjects with JS/MKS and
Jeune syndrome. Finally, we showed that the defect in GCP proliferation was similar in the cerebellar vermis and hemispheres in all patients with ciliopathy analyzed, suggesting that the specific cause of vermal hypo-/aplasia precedes this defect. Our results, obtained from the analysis of human samples, showthat the hemispheres and the vermis are affected in JS/MKS and provide evidence of a defective cellular mechanism in these pathologic processes.

Proc Natl Acad Sci U S A. 2012 Oct 1. [Epub ahead of print]

Features of the Arabidopsis recombination landscape resulting from the combined loss of sequence variation and DNA methylation

Marie Embs — 2015-02-25T10:45:12Z

The rate of meiotic crossing over (CO) varies considerably along chromosomes, leading to marked distortions between physical and genetic distances. The causes underlying this variation are being unraveled, and DNA sequence and chromatin states have emerged as key factors. However, the extent to which the suppression of COs within the repeat-rich pericentromeric regions of plant and mammalian chromosomes results from their high level of DNA polymorphisms and from their heterochromatic state, notably their dense DNA methylation, remains unknown. Here, we test the combined effect of removing sequence polymorphisms and repeat-associated DNA methylation on the meiotic recombination landscape of an Arabidopsis mapping population. To do so, we use genome-wide DNA methylation data from a large panel of isogenic epigenetic recombinant inbred lines (epiRILs) to derive a recombination map based on 126 meiotically stable, differentially methylated regions covering 81.9% of the genome. We demonstrate that the suppression of COs within pericentromeric regions of chromosomes persists in this experimental setting. Moreover, suppression is reinforced within 3-Mb regions flanking pericentromeric boundaries, and this effect appears to be compensated by increased recombination activity in chromosome arms. A direct comparison with 17 classical Arabidopsis crosses shows that these recombination changes place the epiRILs at the boundary of the range of natural variation but are not severe enough to transgress that boundary significantly. This level of robustness is remarkable, considering that this population represents an extreme with key recombination barriers having been forced to a minimum.

Published online before print September 17, 2012, doi : 10.1073/pnas.1212955109

Single-molecule mechanical identification and sequencing

Marie Embs — 2015-02-25T10:43:40Z

High-throughput, low-cost DNA sequencing has emerged as one of the challenges of the postgenomic era. Here we present the proof of concept for a single-molecule platform that allows DNA identification and sequencing. In contrast to most present methods, our scheme is not based on the detection of the fluorescent nucleotides but on DNA hairpin length. By pulling on magnetic beads tethered by a DNA hairpin to the surface, the molecule can be unzipped. In this open state it can hybridize with complementary oligonucleotides, which transiently block the hairpin rezipping when the pulling force is reduced. By measuring from the surface to the bead of a blocked hairpin, one can determine the position of the hybrid along the molecule with nearly single-base precision. Our approach can be used to identify a DNA fragment of known sequence in a mix of various fragments and to sequence an unknown DNA fragment by hybridization or ligation.

Nature Methods (2012) doi:10.1038/nmeth.1925

Polymerase Exchange during Okazaki Fragment Synthesis observed in Living Cells

Marie Embs — 2012-12-01T10:11:00Z

Despite the large amount of work dedicated to its study, DNA replication is still not fully understood.
Recently a group from Oxford university has demonstrated that in E. coli a replication fork contains 3 polymerases and not 2 as it was expected since a priori there is only one needed for each of the 2 DNA strands.
The role of this supplementary polymerase was unknown.

In a collaborative work between a group from the CGM in Gif and a group from the IBENS and the Laboratoire de physique statistique de l'ENS, researchers used a single molecule detection technique to monitor in real time the protein composition of the replisome in a living E. coli. The authors could directly observe that at least one polymerase from the replisome is regularly exchanged, typically every second, with one diffusing in the cellular medium.

By observing in real time the amount of Single Strand Binding proteins at the replication fork, the authors noticed that this polymerase exchange is strongly correlated with the Okazaki fragment synthesis. They could then deduce that the exchanged polymerase is involved in the lagging strand synthesis.
This observation led them to propose that the third polymerase is likely present at the fork to replace the lagging strand one in cases where the capture of a new polymerase from the cellular medium is delayed. This "replacement" polymerase would be for DNA replication like a spare wheel for your car.

Science, 335, 328 (2012), DOI : 10.1126/science.1210400